Directed Evolution of AAV Targeting Primate Retina by Intravitreal Injection Identifies R100, a Variant Demonstrating Robust Gene Delivery and Therapeutic Efficacy in Non-Human Primates
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Targeted AAV vectors are needed for safe and efficient delivery to and transduction of specific tissue target(s) in patients. Effective intravitreal delivery for retina gene therapy is not feasible with wildtype AAV. We employed directed evolution in nonhuman primates (NHP) to discover an AAV variant (R100) for intravitreal treatment of multiple target cells in the primate retina. R100 demonstrated superior transduction of human retinal cells compared to wildtype AAV. Furthermore, three R100-based gene therapeutics demonstrated safety, delivery, and durable pan-retinal expression of intracellular or secreted transgenes throughout the NHP retina following intravitreal administration. Finally, efficacy of R100-mediated delivery of therapeutic transgenes was demonstrated in patient-derived retinal cells (monogenic diseases) and in an NHP model of pathogenic retinal angiogenesis.
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The R100 capsid is capable of improved ILM penetration following intravitreal administration.
“Another 'evolved ' AAV capsid capable of improved ILM penetration following intravitreal administration is the R100 capsid from 4D Mo lecular Therapeutics (Emeryville, CA, USA), discovered using directed evolution in NHPs (Kotterman et al., 2021).”
From Gene Therapy-Associated Uveitis (GTAU): Understanding and mitigating the adverse immune response in retinal gene therapy (2025) · cites this paper as Kotterman et al., 2021
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Cited by
Sorted by citations
Optogenetic Therapy for Visual Restoration
Sakai, D., Tomita, H., & Maeda, A. (2022) · International Journal of Molecular Sciences · doi:10.3390/ijms232315041 · Cited by 62 · see citing works
Adeno-Associated Virus (AAV) - Based Gene Therapies for Retinal Diseases: Where are We?
Ail, D., Malki, H., Zin, E. A., & Dalkara, D. (2023) · The Application of Clinical Genetics · doi:10.2147/tacg.s383453 · Cited by 40 · see citing works
Castro, B. F. M., Steel, J. C., & Layton, C. J. (2023) · BioDrugs · doi:10.1007/s40259-023-00629-y · Cited by 39 · see citing works
Gene Therapies in Clinical Development to Treat Retinal Disorders
McClements, M. E., Elsayed, M. E. A. A., Major, L., de la Cámara, C. M.-F., & MacLaren, R. E. (2024) · Molecular Diagnosis & Therapy · doi:10.1007/s40291-024-00722-0 · Cited by 28 · see citing works
Purdy, R., John, M., Bray, A., Clare, A. J., Copland, D. A., Chan, Y. K., Henderson, R. H., Nerinckx, F., Leroy, B. P., Yang, P., et al. (2025) · Progress in Retinal and Eye Research · doi:10.1016/j.preteyeres.2025.101354 · Cited by 24 · see citing works
Retina-directed gene therapy: Achievements and remaining challenges
Biber, J., Gandor, C. L. A., Bećirović, E., & Michalakis, S. (2025) · Pharmacology & Therapeutics · doi:10.1016/j.pharmthera.2025.108862 · Cited by 24 · see citing works
Gene therapy for age-related macular degeneration: a promising frontier in vision preservation
Hushmandi, K., Lam, H. Y., Wong, W., Tan, W., Daryabari, S., Reiter, R. J., Farahani, N., & Kumar, A. P. (2025) · Cell Communication and Signaling · doi:10.1186/s12964-025-02246-4 · Cited by 21 · see citing works
Adeno-associated virus vectors for retinal gene therapy in basic research and clinical studies
Xia, X., & Guo, X. V. (2023) · Frontiers in Medicine · doi:10.3389/fmed.2023.1310050 · Cited by 20 · see citing works
Christopher, M. A., Hoseini, P., Walker, E., Proudfoot, J. A., Bowd, C., Fazio, M. A., Girkin, C. A., Moraes, C. G. D., Liebmann, J. M., Weinreb, R. N., et al. (2022) · Ophthalmology Glaucoma · doi:10.1016/j.ogla.2022.08.014 · Cited by 12 · see citing works
Clerck, K. D., Smedt, S. C. D., Remaut, K., & Peynshaert, K. (2025) · Journal of Controlled Release · doi:10.1016/j.jconrel.2025.113849 · Cited by 11 · see citing works
Developing New Vectors for Retinal Gene TherapyPreprint
Zin, E. A., Öztürk, B. E., Dalkara, D., & Byrne, L. C. (2023) · Cold Spring Harbor Perspectives in Medicine · doi:10.1101/cshperspect.a041291 · Cited by 12 · see citing works
Shim, J., Kim, Y., Bak, J., Shin, S. H., Lee, K., Hwang, Y. H., Kong, H. Y., & Han, J. S. (2024) · Molecular Therapy — Methods & Clinical Development · doi:10.1016/j.omtm.2024.101366 · Cited by 7 · see citing works
Tuli, N., Moroz, H., Jaffer, A., Kulbay, M., Tanya, S. M., Aslan, F. Ş., Ozdemir, D., Ali, S. K., & Qian, C. X. (2026) · Diagnostics · doi:10.3390/diagnostics16030392 · Cited by 4 · see citing works
AAV-based gene therapies for neovascular AMD
Kim, T. H., Kwon, C. Y., Song, J. Y., & Yoo, H. C. (2026) · Gene Therapy · doi:10.1038/s41434-026-00595-4 · Cited by 4 · see citing works
Luo, L., Xu, J., Wang, B.-Q., Chen, C., Chen, X., Hu, Q., Wang, Y., Zhang, W.-Y., Jiang, W., Li, X.-T., et al. (2023) · Biomaterials · doi:10.1016/j.biomaterials.2023.122403 · Cited by 1 · see citing works
Structural dynamics insights into principles underlying the fitness of new broadly potent AAVsPreprint
Johnson, M. E., Öztürk, B. E., Tugwell, T. H., Lambros, M., Janowitz, H. N., Flohr, K., Sedorovitz, M., Campello, L., Hartung, J., Hogle, B. C., et al. (2026) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.64898/2026.03.24.713814 · Cited by 1 · see citing works
Recent Development in in vivo Clinical Gene Therapy Platforms
Murphy, J. E., & Owens, J. M. (2024) · Wiley eBooks · doi:10.1002/9781119852810.ch2
Viability of engineered AAVs via protein language modelsPreprint
Desrosiers, M., Ocari, T., Trinquier, J., Zin, E. A., Meter, T. V., Delmas, M., Tekinsoy, M., Planul, A., Nemoto, T., Dalkara, D., et al. (2026) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.64898/2026.06.11.731521
- Authors
- Kotterman, M., Beliakoff, G., Croze, R., Vazin, T., Schmitt, C., Szymanski, P., Leong, M., Quezada, M., Holt, J., Barglow, K., Hassanipour, M., Schaffer, D., Francis, P., & Kirn, D.
- Affiliations
- University of California, Berkeley (US)
- Corresponding author
- Kotterman, Melissa
- Venue
- bioRxiv (Cold Spring Harbor Laboratory)
- Publisher
- openRxiv
- Published
- 2021-06-25
- OpenAlex
- W3176322268
- Cited by
- 20
- References
- 87
- Citations by year
- 2026: 4 · 2025: 4 · 2024: 3 · 2023: 5 · 2022: 3 · 2021: 1
- Type
- posted content
- Language
- en
- Open access
- green
- doi.org
- Locations
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
- Funders
- F. Hoffmann-La Roche
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Retinal Development and Disorders
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Keywords
- adeno-associated virus, intravitreal injection, retinal gene therapy, primate retina, non-human primates, retinal transduction, retinal angiogenesis, gene delivery
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
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